Colorectal carcinoma is the second most frequent cause of cancer mortality in men and women, causing nearly one third of all malignancy-related deaths in North America. It has been estimated that ultimately as many as 6% of Canadians and Americans will develop malignancy in the lower bowel, and over 50% of them will die within 5 years of diagnosis. Because there are no realistic prospects of significantly improving the cure rate once the cancer has spread beyond the bowel wall, many authorities believe that colorectal cancer can be controlled only by preventive measures (1).
Primary prevention, i.e. averting the development of the tumour by altering biological risk factors, is not yet feasible since so little is understood of the etiology of the disease. Alternatively, secondary preventive measures, i.e. detection at an asymptomatic, treatable state, would be possible should an effective screening test be available. Indeed, neoplasms of the lower bowel have the characteristics that make them a suitable candidate for the development of a screening test. This is because i) it is a common cause of cancer-related deaths, and ii) whereas once the stage of true cancer is reached, leading to symptoms, the mortality rate is over 50%, removal of bowel neoplasms at its earliest, asymptomatic stage can be done by non-surgical endoscopic polypectomy, without any significant risk. Moreover, it requires at least four to six years before an adenomatous polyp reaches the cancer stage, so there is ample opportunity to detect these neoplasms at their treatable stage. Recent clinical studies document a decrease in mortality from colorectal cancer screening, as predicted by these theoretical considerations. The problem to-date has been that polyps can be reliably detected only by endoscopy.
Thus, colorectal cancer satisfies each of the following three criteria of a disease considered suitable for a screening program. First, it is a relatively common condition with serious consequences. Second, curative treatment is available when detected at an early stage, i.e. snare polypectomy through a colonoscope or surgical segmental bowel resection. Third, the prevalence is sufficiently high to justify the expense of a screening program (2).
PRINCIPLES OF SCREENING
The goal of a medical screening program is to reduce morbidity and mortality by detecting a disease at a sufficiently early stage to allow curative treatment. It is not designed necessarily to diagnose a disease, but to determine which asymptomatic, apparently disease-free individuals should undergo diagnostic investigations. The ability of a screening test to distinguish those who warrant further evaluation from those who do not is expressed in epidemiological terms. The term "sensitivity" is defined as the proportion of diseased individuals who have a positive test, i.e. the proportion of true positives/all persons with the disease. "Specificity" is the proportion of disease-free subjects who have a negative test, i.e. the proportion of true negatives/all persons without the disease. The term "positive predictive value" is the proportion of positive tests due to the disease, i.e. the proportion of true positives/all positives. Almost always, sensitivity and specificity must be traded against each another. Intuitively, it appears wise to design a screening test for a fatal disease so as to optimize sensitivity, in order to detect as many individuals with the disease as possible. It has been emphasized, however, that optimizing sensitivity brings with it a risk of reducing specificity to such an extent that unacceptably high costs, poor compliance, and "flooding" of diagnostic facilities result. Moreover, positive predictive value, which is a particularly useful expression of the value of a screening test, is critically dependent on specificity and on the prevalence of the disease in the population screened.
It has been stressed that the effectiveness of a screening test can be properly evaluated only by randomized controlled trials. In the case of cancer, it is not sufficient to demonstrate that life is prolonged when the malignancy is detected by a positive screening test, compared to when the tumour is diagnosed after the development of symptoms. Instead, it must be shown that screened individuals have a lower death rate from the malignancy than similar individuals not enrolled in such a screening program. Important sources of error in interpreting the results of previous screening programs include lead time bias, length bias, and patient selection bias. A particularly fallacious assumption is that the predictive value of a screening test is the same in a hospitalized population with advanced disease, in which the test is usually initially tried, as it is in a healthy population with early minimal disease, to which the test is usually aimed.
CURRENT POPULATION SCREENING METHODS
Endoscopic methods, such as sigmoidoscopy or entire-length colonoscopy, are diagnostic rather than screening techniques. The only current method of colorectal cancer screening in the general population is searching for occult blood in the stool (3). Present techniques e.g. HemOccult II involve smearing a sample of stool onto guaiac impregnated paper which, after treatment with hydrogen peroxide containing developer, exhibits blue colour if blood, haemoglobin, is present. After almost two decades of experience with this methodology, it has become clear that even in expert centres, the sensitivity is less than 50% for curable neoplasms, and that the positive predictive value approximates, at best, only 40% in a clinic population. An update from the large-scale (n=97, 205) University of Minnesota, Minnesota, United States, prospective trial indicates a positive predictive value for colorectal cancer of only 2.2% when HemOccult is used in asymptomatic subjects, aged 50-80, with an overall disease prevalence of 0.2% (4). Furthermore, factors such as medications, multiple dietary constituents, delays in specimen handling, variabilities in fecal hydration, and storage of assay materials commonly confound results. Analysis of one of the three randomized controlled studies assessing the value of HemOccult suggests comparable mortality rates in the screened and control populations (5). Newer methods of detecting occult blood, e.g. methods based either on porphyrin analysis [HemoQuant] or antibody specific for human haemoglobin, may improve on these results. However, three limiting problems remain unlikely to be overcome. These are that colorectal malignancies shed blood only intermittently, upper gastrointestinal tract bleeding may make the results (falsely) positive, and multiple lesions in the lower bowel, apart from colorectal neoplasms, commonly bleed. Such lesions include hemorrhoids, diverticulae, ulcers, and vascular ectasie. Compliance in unselected populations has been estimated to be less than 30%, at least partly because the technique requires patients themselves to smear their stool onto a slide or a strip, a task most people find not only distasteful, but also technically difficult. Despite this, HemOccult continues to be widely used because the American Cancer Society has recommended occult blood testing yearly for all individuals over 50 years of age, arguing that even an imperfect test will save many lives. Implicit in all arguments over the value of HemOccuIt is that any improvement in screening techniques for bowel malignancy would have a dramatic impact on colorectal cancer mortality rates from the disease, since the screening for occult blood even in the present form leads to reducing mortality from colorectal cancer (6) .
EXPERIMENTAL SCREENING METHODS (i) Screening for colorectal cancer by stool DNA analysis (7).
This is based on the presence in stool of neoplastic cells shed in large numbers into the colonic lumen. In principle, a mutation which is common to neoplasms could be detected with high precision by analyzing DNA from these cells. Therefore, the existence of a detectable mutation in the colorectal tumour is a prerequisite for developing such a method of screening. Unfortunately, this technique can recognize a mutation based only on a new or altered oligonucleotide sequence, but not on a loss of its portion. Thus, neoplasia-related mutations based on deletion in genes, e.g. allele losses on chromosomes such as are commonly found in colorectal tumours, are beyond the limits of the methodology. Currently, the most common mutation is the K-ras oncogene mutation present, in about 40% of colorectal carcinomas and adenomas. Screening for K-ras gene can therefore detect, at best, only 40% of all neoplasias. This methodology is at present technically very complex and expensive. (ii) Screening for the presence in colonic mucin of a cancer-related disaccharide, D-Galp(.beta.1-3)-D-GalpNAc(.alpha.l,Ser/Thr), T-(Thomsen-Friedenreich) antigen, since it is widely known that T-antigen is not expressed by cells in healthy colons, whereas it is expressed by cancer (8). (a) Monoclonal antibodies and lectins: It has been shown that monoclonal antibodies raised against synthetic T-antigen recognize and bind to cancer cells. Similarly, peanut agglutinin (PNA), a lectin, binds strongly to the same disaccharide, but recognizes malignancy with lesser specificity. Amaranthin, a lectin from Amaranthus caudatus, has been reported to have better specificity for T-antigen than PNA. Neither amaranthin nor PNA bind to histological sections of normal mucosa, but both bind to mucin in the goblet cells of tumours and certain polyps, and in the transitional mucosa. The visualization of the binding utilizes fluorescently labelled antibodies and lectins (9). (b) Galactose oxidase test. T-antigen is also reported to be detectable colorimetrically after oxidation of OH-6 of galactose using galactose oxidase and visualization of the resulting aldehyde with Schiff's reagent, - U.S. Pat. No. 4,857,457, issued Aug. 15, 1989 to Shamsuddin et al. In contrast with the tests using lectins, this test is performed on mucus samples obtained by digital rectal examination and smeared onto a support. This system demonstrated a sensitivity of 74% and specificity of 50% for colorectal neoplasms, i.e. adenomatous polyps and cancer, in one study with only 1 false negative result among 59 patients with cancer. Since then a number of reports of basically the same test have appeared with sensitivity ranging from 35% to 100% and specificity ranging from 15% to 76%. Some investigators found that the test was more sensitive, but less specific, than HemOccult. The lesser specificity has been ascribed to the positivity of test in individuals with certain inflammatory condition, such as diverticulitis and ulcerative colitis (10).